1414
6, 1977)
Acid Digestion, Hydride Evolution Atomic Absorption Spectro-
photometric Method for Determining Arsenic and Selenium in
Foods: Collaborative Study. Part I
MILAN IHNAT and HANFORD J. MILLER1
Agriculture Canada, Chemistry and Biology Research Institute, Ottawa, Ontario, Canada KlA 0C6
The hydride evolution atomic absorption spec-
trophotometric (AAS) method for determining
As and Se in foods developed and evaluated by
the Food and Drug Administration and Agri-
culture Canada laboratories was subjected to
collaborative study. Twenty-four laboratories
provided results for As and 23 provided results
for Se levels in 13 different samples consisting
of tuna, swordfish, flounder, oyster, liver, flour,
skim milk powder, spinach, kale, and apple con-
taining natural levels of As and Se in the ranges
0-15,000 and 0-4000 ng/g, respectively. Ref-
erence materials formed a substantial segment
of samples, and a number of other laboratories
using fluorometry, colorimetry, neutron activa-
tion, spark source mass spectrometry, and
graphite furnace AAS provided confirmative ref-
erence values for the remaining samples. A
variety of hydride generation instruments were
used, ranging from commercially available de-
vices to semiautomated and fully automated
custom-made instruments. Although the accu-
racy of the method was fairly good, between-
laboratory and between-determination (hydride
evolution AAS measurement) precisions were
not favorable. The main advantage of the hy-
dride AAS method is the rapidity of the deter-
minative step.
The current official methods (1) for deter-
mining arsenic and selenium in foods are based
on arsine generation-light absorption spectro-
photometry (colorimetry) for arsenic (25.006-
25.019) and fluorometry for selenium (25.117-
25.120), replacing the Gutzeit method for ar-
senic (25.020) and the thiosulfate volumetric
method for selenium (25.121-25.126), both of
which are now surplus methods. The fluoro-
metric method for selenium has a detection limit
of about 10-20 ng, but the arsenic methods lack
the capability for estimating arsenic at nano-
gram levels.
1 2
Food and Drug Administration, 900 Madison Ave, Balti-
more, MD 21201. Present address: U.S. Geological Survey,
Water Resources Division, Box 25046, Denver Federal Center,
Denver, CO 80225.
JOURNAL OF THE AOAC (Vol. 60, N o .
Advantages of analytical procedures based on
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hydride evolution atomic absorption spectropho-
tometry (AAS) include nanogram detection lim-
its, a rapid determinative step,2 and capacity for
simultaneous determination of both elements.
The acid digestion, hydride evolution AAS
method for determining arsenic and selenium in
foods developed and evaluated by the Food and
Drug Administration (FDA) and Agriculture
Canada laboratories (2) was subjected to a col-
laborative study; the results are presented in
this and following reports. A description of the
study and the overall performance of the
method are reported here while the following
papers (3, 4) deal with a detailed treatment and
statistical analysis of the results; all reports are
integral to the assessment of the method.
Collaborative Study
Description of Samples
Thirteen samples representing 6 major classes
of food (fruit, vegetables, cereal, dairy products,
meat, and fish) were selected to evaluate the per-
formance of the method. The samples, sample and
analytical solution code numbers, moisture con-
tents, and weights requested to be taken for analy-
sis are listed in Table 1. They were grouped to
provide each laboratory with 14 samples (12 dif-
ferent samples) for analysis; half the collaborators
received Sample 5A, the other half received Sam-
ple 5B. The shortage of some samples, particularly
expensive reference materials, usually required
sending the minimum amounts necessary for the
analyses requested. Shortages of orchard leaves
(Sample 1), tuna (Sample 2), bovine liver (Sam-
ple 8), and spinach (Sample 9) resulted in either
randomly excluding some samples or providing
amounts sufficient for only one analysis.
Apples, skim milk powder, and flour were pur-
chased from local retail outlets; flounder and
swordfish samples were obtained from Environ-
ment Canada. Reference materials constituted a
Determinative step and determination refer to the hydride
generation AAS measurement step (i.e., digest -> result);
analysis and method refer to the entire analytical procedure
(i.e., sample -> result).
IHNAT & MILLER: HYDRIDE AAS METHOD FOR AS AND SE IN FOOD 1415

Table 1. Description of samples used in collaborative study

Sample Moisture Wt analyzed, Anal.
codes" Description 6 content, %' Ed soln code'

Practice flounder 2.1 _

Pi. P 2 .. •
1 1 NBS Orchard Leaves 6.8 2.000, 2.000 It, 1 2
2 A NBS Tuna 3.0 1.000, 1.000 2i, 22
3 B IAEA oyster 6.7 1.000 3
4 C swordfish-' 2.1 1.000 4
5A JA apple (Macintosh)" 3.8 1.000 5A
5B JB apple (Delicious)' 5.9 1.000 5B

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6 E flounder 2.1 1.000 6
7 H kale 7.9 2.000 7
8 D NBS Bovine Liver 6.2 2.000, 2.000 8|, 82
9 K NBS Spinach 4.6 2.000, 2.000 9|, 92
10 F IAEA flour 10.8 2.000 10
11 L skim milk powder' 4.9 2.000 11
12 G flour' 9.6 2.000 12
13 B IAEA oyster 6.7 1.000 13
14 H kale 7.9 2.000 14

Standard Solution Ampoules

A As and Se A,, A 2
B As and Se Bi, B2
C As and Se C,, C2
D As* —

" First column is collaborative laboratory code; second column is reference laboratory code.
6
All samples were dry or freeze-dried materials. The 2 flounder samples, 2 oyster samples, and 2 kale samples
were blind duplicates.
c
All samples except 1, 7, and 14 were dried under vacuum 15 hr at 70°C; Samples 1, 7, and 14 were dried under
vacuum 24 hr at 90°C. Mean of 2-5 analyses.
* Weight requested; actual weights used differed in some cases.
• Subscripts 1 and 2 for samples and ampoule C refer to first and second digestions, respectively. Ai, B|, A2,
and B2 refer to undigested solutions prepared from ampoule contents.
1
The swordfish, skim milk powder, and flour samples were identical to those used in a previous collaborative
study (5) where they were numbered 8, 2, and 9, respectively.
" One half of the collaborators received Sample 5A, the other half received Sample 5B.
h
Used for spiking Samples 5A and 5B.

substantial segment of samples and included NBS
Standard Reference Materials Spinach (SRM
1570), Orchard Leaves (SRM 1571), Bovine Liver
(SRM 1577) and Tuna (not yet issued); Inter-
national Atomic Energy Agency (IAEA) oyster
(MA-M-1) and flour (V-2/1 (1974)); and kale
(H. J. M. Bowen). In addition, 3 samples, sword-
fish (Sample 4), skim milk powder (Sample 11),
and flour (Sample 12) were used in the 1973 col-
laborative study of the fluorometric method for
Se (5); extensive analytical data concerning Se
were available for these samples.
For ease of handling, all samples were shipped
in screw-capped glass vials as either dry or freeze-
dried materials. Flounder, swordfish, and apple
samples were prepared by homogenizing in a
blender, freeze-drying, grinding, and mixing; the
skim milk powder was ground to a fine powder;
the NBS and IAEA samples and kale were used
as received. During packaging, 2-5 vials of each
sample were set aside for determination of mois-
ture; it ranged from 2.1 to 10.8% in the different
samples. Samples contained levels of native As
and Se in the ranges of 0-15,000 and 0-4000 ng/g,
respectively. Only the apple samples (Samples 5A
and 5B) were fortified with As by collaborators,
because native levels were low and we wanted to
evaluate the performance of the method for this
food matrix.
Four glass ampoules containing standard solu-
tions of inorganic As (III) and Se (IV) in IN
H2SO4 at concentrations of 625 ng As + 625 ng
Se/ml (ampoule A), 1250 ng A s + 1250 ng Se/ml
(ampoules B and C), and 1250 ng As/ml (ampoule
D) were included to monitor instrument perform-
ance and the digestion step (ampoules A, B, and
C), and to provide a solution of As for fortifying
Samples 5A and 5B (ampoule D).
Description of Study
Each collaborating laboratory was supplied with
the 14 samples (unidentified), and 1 practice sam-
ple (flounder containing 15,000 ng As and 1500 ng
Se/g). Collaborators supplied their own reagents
and equipment, and were instructed to request
elemental Se for preparing standard solutions if
1416
that reagent was unavailable. A copy of the method
(2), additional instructions, and report forms were
included.
The study was designed to amass a large body
of information in an attempt to relate the per-
formance of the sample treatment and determina-
tive steps to a number of variables. Parameters
considered were analyst experience, type of hy-
dride generation apparatus and technique, repro-
ducibility and type of calibration curves, instru-
mental baseline noise, analytical precision and
accuracy, detection limits and sensitivities of the
determinative step and the method, absorbance-
time peak shape and reproducibility, digestion
technique, digestion and determination sequences,
recovery of inorganic As and Se from untreated
standard solutions and digested standard solutions
and fortified samples, undigested and digested
reagent blank magnitudes, variabilities and origins,
sample type, and analyte levels. Consequently, a
large number of analyses/determinations were re-
quested from each laboratory, together with de-
tailed information regarding the experimental
technique. The raw experimental data in the form
of recorder charts were also requested. Many labo-
ratories were invited so that a number of different
commercial, modified, and custom-built hydride
generators would be included and sufficient num-
bers of collaborators would use the same types to
permit statistical assessment of several specific
types of generators. Another, smaller group of
laboratories, denoted reference laboratories, was
approached to provide independent analyses of the
samples, particularly the non-certified samples,
using analytical techniques other than hydride evo-
lution AAS. It was expected that these results
together with certified reference and other data
would give good estimates of As and Se concen-
trations for evaluating the hydride method ac-
curacy. Manufacturers of hydride generation in-
strumentation and AAS equipment were con-
tacted for relevant information on operating de-
tails and performance characteristics of, in particu-
lar, the former, so that this information could be
on hand when collaborative results were received.
The scheme of sample coding and the analyses
requested gave 6 pairs of duplicate results, and 7
unpaired sets of data. Samples 3 and 13 as well
as Samples 7 and 14 were, unknown to the col-
laborators, identical samples of IAEA oyster and
Bowen's kale, respectively, serving as blind dupli-
cates and augmenting the 4 known pairs (lj, 12),
(2lF 22), (8j, 82), and (9X, 9 2 ). In addition Sample
6 was identical to the practice sample which could
Contribution No. 961 from the Chemistry and Biology
Research Institute.
JOURNAL OF THE AOAC (Vol. 60, No. 6, 1977)
provide a possible third pair of blind duplicates
should practice results be reported and be accept-
able. Ampoule solutions A, B, and C were pre-
pared to contain equal concentrations of As and Se
so that, when diluted, concentrations would be Al5
10.0 ng/ml; A2, 1.0 ng/ml; B ^ Ci, and C 2 , 20.0
ng/ml; and B 2 , 2.0 ng/ml. Incorporation of these
standard solutions was expected to provide infor-
mation on detectivity, accuracy, and systematic
error of the determinative step, and effect of
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digestion on analytes in simple matrices.
Collaborators were requested to follow specific
sample digestion and determination sequences.
Laboratories were randomly assigned 1 of the 8
permutations of sequences. This approach was
adopted in an attempt to isolate any systematic
error due to differences in performance of collabo-
rators as they became more familiar with the pro-
cedure. Samples were grouped according to analyte
levels to facilitate AAS measurements; for those
samples with moderate to high concentrations, an
instrumental scale expansion of Xl was recom-
mended; a higher scale expansion was suggested
for low level samples. The magnitude of the higher
scale expansion was left to the discretion of the
analyst because it depends on the performance of
individual instrumentation. It was requested that
all determinations be conducted in duplicate to
enable evaluation of this step of the method.
The greatest difficulty in the design of the study
was related to the wide range of hydride generators
in use with divergent detectivities, solution volume
requirements (1-100 ml), and other performance
characteristics (e.g., standard curve linearity), and
the desire to make the experiment suitable to all
types of generators used by collaborators. As a
compromise, the digests of 1 g high analyte level
samples or 2 g low level samples were diluted to
100 ml, and collaborators were requested to use
aliquots suitable to their particular instrumenta-
tion, diluting if necessary to get absorbance read-
ings on scale.
Collaborators were instructed that the digestion
procedure was firmly specified and should be fol-
lowed closely. Other details of the method, how-
ever, particularly those related to the hydride gen-
erator, the acid and reductant concentrations, and
the measurement step, were left to the laboratories
to decide and implement appropriate procedures
for their particular instrumentation. Samples
were assembled in 4 groups of 6 so collaborators
could use one or more 6-element digestors, 12-
element digestors, or whatever their capability al-
lowed. They were instructed to use the practice
sample provided to gain experience with the
method and to ensure that everything was func-
tioning normally. All samples were dry powders,
IHNAT & MILLER: HYDRIDE AAS METHOD FOR AS AND SE IN FOOD
and were not to be further dried or exposed longer
than necessary to complete the weighing.
The ampoules contained standards in acid solu-
tion. Working solutions were to be prepared as
follows: Pipet 4.00 ml solution from ampoule A
into 250 ml volumetric flask and dilute to volume
with the 5% H 2 SO 4 + 30% HC1 diluting solution,
or similar solution geared to hydride apparatus to
give working solution Ax. Dilute 25 ml solution Ax
to 250 ml with the same acid mixture to give
solution A2. In like manner, prepare solutions Bj
and B 2 from ampoule B. Pipet 4.00 ml of solution
from ampoule C into 25 ml volumetric flask and
dilute to volume with water to give solution C;
duplicate 10 ml digests diluted to 100 ml are de-
noted solutions Ci and C2. Prepare solution D by
similarly diluting 4.00 ml of the contents of am-
poule D to 25 ml with water. Add 5 ml solution D
to 1.00 g Sample 5 in Kjeldahl flask to yield forti-
fied sample.
Rectify charred samples as indicated, salvaging
sample if possible. Report results for both the
charred sample and a repeat sample if possible.
Pipet 10.0 ml solution C into Kjeldahl flask, add
digestion acid mixture, and proceed with digestion.
Repeat process with second 10.0 ml aliquot. For
Sample 5A or 5B, weigh 1.00 g sample into Kjel-
dahl flask, pipet in 5.00 ml solution D, add acid
digestion mixture, and proceed with digestion.
Ensure that each standard (calibration) curve
contains 5 points, including reagent blank, cover-
ing range of instrumentation at each scale expan-
sion. Prepare all standard solutions from Se° and
As2O3 (commercial standard solutions may be
used). Standard solutions and associated reagent
blanks are not digested; all samples, solution C,
and associated reagent blanks are digested.
Use whatever solution volume is required by
generator, taking up to 20 ml aliquot from sample
solution and making any dilutions to required vol-
ume with H2SO4-HC1 mixture. If NaBH 4 and
H2SO.|-HC1 concentrations specified are unsuitable,
use concentrations appropriate to specific situa-
tion. Try to use same acid matrices for both As
and Se, and adjust acid content with HC1 because
5 ml H 2 SO 4 remains after digestion procedure
which is firmly specified.
When samples give off-scale absorbance readings,
repeat analyses, using <%th sample solution
volume. It may be necessary to dilute solutions
for automated equipment.
In determination sequence (hydride generation
and measurement), use the standard solution and
reagent blank absorbance readings preceding the
sample readings in each run for calculating As
and Se concentrations for that run only. The entire
determination consists of 18 runs, 8 for each ele-
1417
ment to determine concentrations, and one for
each element (at a high recorder chart-speed, at
least 150-200 mm/min) to provide information on
peak shape. Fit breaks in the determination at the
end of a run. A long break, particularly if flame is
extinguished, may warrant a recheck of instrument
parameters. If more than one day is necessary to
complete the determination, carry out one-half of
the determinations on each of 2 days. Leave chart
running continuously during all the determina-
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tions, except during high speed determinations, to
provide information on noise, drift, etc., under con-
stant instrument settings. For high chart speed
runs, record only the peaks, but include sufficient
baseline before and after the peak.
Collaborators were requested to submit the
original high chart speed tracings, and, if possible,
the entire original chart record with appropriate
notations of parameters and analysis conditions.
Collaborators were requested to report absorbances
of the undigested reagent blanks and the standard
solutions for all analyses and to indicate whether
an actual absorbance or other instrument mode
was used. Concentrations of As and Se (or at least
absorbances, if concentrations were not calculated),
found in the 4 digested reagent blanks RB^ RB 2 ,
RB 3 , and RB 4 and volumes used for hydride gen-
eration were requested. The nature and use of the
2 types of blanks were explained diagrammatically.
For all samples and ampoule solutions it was re-
quested that aliquots of solution taken for hydride
generation, analyte concentrations, and contents
in the aliquots be reported; in addition, for sam-
ples, analyte concentrations (ng/g) were requested.
Information on the following instrumental and
flame parameters was requested: atomic absorp-
tion spectrophotometer and recorder makes and
models, resonance line wavelength, monochromator
slit width and spectral bandpass, scale expansion,
signal damping, lamp type and operating current
or wattage, recorder input voltage for full scale
deflection, chart speed for routine and high speed
runs, flame gases and flow rates, burner head type
and slot dimensions, and height of center of light
beam above burner. Inquiries were also made re-
garding the use of draft shielding for the flame,
whether spectrometer parameters were optimized
using criteria of sensitivities (highest signal) or
detection limits (highest signal/noise), and whether
standard solutions or hydride generation was used
for optimization. The following information con-
cerning the hydride generation apparatus was so-
licited: make and model of the generator, and
whether commercial, modified, or custom-made;
volume of solution required by the generator and
actual volume used in this study; acid mixture
used in this study, whether it differed from the
1418
collaborator's normal operations, and whether the
concentration was optimum for both arsenic and
selenium determinations; nature and form of re-
ductant (NaBH 4 was specified in method); re-
ductant solution concentration and composition
(e.g., NaOH stabilizer concentration); amount or
volume of reductant used per determination; and
type of auxiliary carrier gas and flow rate. Col-
laborators were asked to describe operating in-
structions and construction details if they differed
from those for commercial apparatus or if custom
designs were used. It was emphasized that volumes
and operating conditions optimum for specific ap-
paratus used should take precedence over instruc-
tions provided by the initiating laboratories. Other
information requested dealt with sample digestion
details (charring, foaming, etc.), and sources and
purities of NaBH 4 and other reagents. Collabo-
rators were also asked to indicate the experience
of the participating analysts with the wet diges-
tion and hydride generation.
Reference laboratories were free to use analyti-
cal methods with which they had had experience
and in which they had confidence. They were
asked to conduct any number of analyses/deter-
minations for As and Se on any number of samples
provided, and to submit only the data with which
they were satisfied. Sample identities were not
revealed, but ranges of As and Se present (0-0.1,
0.1-1, 1-10 jtig/g) were given to facilitate analysis.
Samples were coded differently than for collabo-
rative laboratories in an attempt to preserve sam-
ple anonymity in instances where the laboratory
was requested to serve the dual role of collabo-
rator and reference laboratory. Reference labora-
tories were requested to report mean As and Se
levels found, standard errors or confidence limits,
and number of analyses/determinations conducted.
They were also asked to report on the analytical
method used, together with literature references
and details and to mention any problems en-
countered.
Results and Discussion
Of the 55 laboratories that agreed to partici-
pate in this study either as collaborators or ref-
erence laboratories, 24 submitted collaborative
analytical results on arsenic and 23 submitted
results on selenium. One collaborator who did
not have HC104 digestion facilities, analyzed
only the undigested ampoule solutions. Two
others used Zn or Al as reductants rather than
NaBH4; their results are presented separately
with data from other laboratories which used
different sample treatment steps (3). Four labo-
ratories attempted analyses but obtained results
JOURNAL OF THE AOAC (Vol. 60, No. 6, 1977)
unsatisfactory to themselves and hence reported
no results. In addition, 11 laboratories reported
reference analyses for methods other than hy-
dride generation.
All hydride generation AAS results for sam-
ples are presented for arsenic and selenium in
Tables 2 and 3, respectively. Although most col-
laborators conducted duplicate determinations as
requested, some reported only averages and not
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the individual results. The submission of raw ex-
perimental data, however, made it possible for
the authors to extract these individual data for
some collaborative sets to provide a larger data
base for calculation of within-digestion variance.
In addition, concentrations could be calculated
in several other instances where results were re-
ported as "not detectable," "less than . . . ," or
"0." These results calculated by the Associate
Referees, are included with reported data in
Tables 2 and 3. When only one of the determi-
nation pair warranted recalculation, levels were
recalculated for both determinations to obviate
artificial inflation of standard deviation due to
incursion of a measurement/calculation bias
likely to be different from that of the collabo-
rator. Calculations were based on raw experi-
mental data provided in the form of recorder
charts, reported peak height measurements, or
reported weights or concentrations of arsenic or
selenium in the solutions and do not necessarily
reflect complete remeasurement/recalculation;
calibration graphs provided by some collabora-
tors were used when deemed suitable. Calculated
concentrations are corrected for digested reagent
blanks in the same determination run as the
sample, regardless of whether the blank was di-
gested with the sample. Negative values are in-
cluded for the purpose of estimating precision
and systematic error. More digits than war-
ranted by the precision of the method have been
retained. Each datum is for one determination
unless otherwise indicated; data include results
extracted, by the Associate Referees, from high
recorder speed runs and other raw data not used
by collaborators. The body of sample concentra-
tion data used for statistical analysis is indi-
cated by underscoring.
The sample concentration data were scanned
for extreme values, and a number of results were
rejected subjectively prior to statistical calcula-
tions for the following reasons: All results for
both elements from Collaborators 3 and 34 were
 Table 2. Collaborative sample results (ng As/g) for the hydride AAS determination of arsenic in foods9
Sample/solution
g
Coll. 2 2 3 4 5A 5B 6 7 8 8 10 n 12 13 14
h h 1 2 i 2 '1 '2
g
t 4(> b 157b
ILL

1 8745 10270^ 2897 3000 11385 3199 1000 - 14080 117 63 150 ND^ ND ND 11180 115
9120 11220^ 2760 3600 12020 2960 880 13980 112 55 38* 162 165^ ND ND ND 12520 115
lp. 12?- 4?-
-p. 4?

50 100 300 <100 120 <100 <50 50 50 <S0 50 <S0 300 50

t
4 9280 9910 1000& 1590 11100 1760 2290& 7830 99 20 24 110 104 <2Ct- <2cr 11260 69
9280 9730 100CT 1460 11100 1710 22OO& 7610 99 20 *2(F- 110 104 28? 29? 7- 9890 69
26? -26?
62?
h h
5 8371h 2773^ 2S94-- 11250^ 2919- 998~ 14881^ 100^- SO* s*e ND| 11239*- 92*
r: HYDRIDE AAS METH

8800 s 8371? 2623? 2614? 11400? 2959? 968-? 14087?- 98? 49? 134? isT? 12s 11239? 92?
8900^ 8596H 2723? 2594? 11650^ 2979^ 1028^ 16667.E. 95? 38? 44? 121? 151- 7? -Z~ T? 10989 s 90?
8450- 28352
O
8b 1605 s 3720 12900 3840 790 14200 40 145 160 150fi 190 11580 65

10 9545 2942 2875 14160 3197 1100 16420 42 30 125 0 12540 65

9245 3115 11040 3047 1020 16500 85 35 168 0 0 0 13720 107
-2| -6? 2-?
O?
o
F
11 106374 3204 10397 1171 16284 236^ 252* 453* 2251 167 11740 171
4677-1 2313 11113 600 1855^ 12712J- 476-L 231& 403^ 465s 117-1 205 11718

12 9540 3128 _ 10509 3643 1240 15684 39 16 146 ND ND ND 11758 251£ 8

8116 2788 10384 3214 1050 14452 36 6 71 ND ND ND 11591 246s
-35? £
-15? 1

13 10000 9350 3230 3000 8660 2670 720 12500 103 40 140 ND ND ND 8000
10250 8750 2270 2000 9500 2600 780 12500 105 38 128 ND ND ND 8500
4?
-i"i? -a?
h h
15* 900CT 10900^ 409CT- 3560*- 12070**- 2970^ 700^ NDe - ND ND ND
7070 s 10160- 4280? 3875- 10730-3 668? 14260^ -38- 33? _29O?>£ -78 s -20? 11910? 100?
9500^ 9780S 3060^ 4010S 11100^ 2790? 2 1462O£ 13870?
2550?
2970(4 2 )-

16 7500- - ND 1800^ ND - -
35(7
i >J -
9500? 750^'* 120^? 285? s. 80^ 100? 218?., -' 168JL •
9500? I 1130? 2630^'^ 655?^a Si 155? 118 i—*

(Contined) CO
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 Table 2. (Continued)
Sample/solution O
Coll. l 2 3 4 5A 5B 6 7 8 8 9 10 ll 12 13 14
h 2 h 2 1 2 '1 2

18* 7700 9100 400s 450| 4200s 600 1060 4700 70 30^ 40 80 90 -40l . -20 5700i 75
8300 7700 600f 60of& 470CT 500 1060 4400 60 30=!- 35 150 120 -40l -20 570C4- 80
8300 9200 7002 850 4600i 900 1100 4500 65 250i 50 110 90 lOOi 50 lOOOoi 65
500i 5200s 680 1100 6560 80 2001 100 110 80 10? 50 97001 70
1070
19 9200 2900 2900 12100 1020 320 0 100 18 130 9 0 0 12900 35
10700 2600 3400 13100 970 370 100 g125 20 160 5 0 3 13900 55
95
f
22 8830*'°. 115002.'° 2960*- 13900*- 102*- 11* 16*^2.116002. 101*
86485 27065 116065 28465 9365 141725 1155 505 1265 255 55 185 121615 95S.
90505 27965 117105 30555 10165 135735 1105 455 1215 205 105 235 120605 1055
23h 9100 8900 4300 4400 12900 4200 2650S. 18900 12600
24* 6700*- - 1980*- 255C& 7102- 109002. - 502- - 85802-
6100£>t 2900i>A 26755 lO6oS 3200^ 530^' 10725- 68s 85 s 25s
s s 1 8675I
53755>i 1050^'i 2525 s 8200 2075s'! 895 s '•i 11175 s 63^ 8 3"? -25- -3- 98^''•"- 850CT 120&J
7300 s 7975s
28 8950 8350 f,a
3610 5060s- 1210 1015ol 195 125. 185 170 85 150 13300 140
8850 3530 470OS- 1270 6150^-'-^- 63001 185 <50— 110 <100- <50^- 60 10800 135

29^ 162004- 6610 s 74304 4310 1350(41 68904 4Ol,s 460 s 4610 s 790s 950s- 0 660s- 0 5 no 1 853O1 35l
1040CH- 3210 s 3610 1 4210 849O 571O 1 300 s 7550s 470s 900s 820s- 240l 158001

30 328C& 12130*. 3010^ - iwsS- 1S42- ND 23825^-

901 if 3760 s 10026s- e U
2943s- 5? 1 32l 164- 10221- 8^
s
87 srf
10213- 2522 s 11623 s 1103 - "63 87^- 137- 11825 s 96 s
JOURNAL OF 1

9412- 12169 s 3020^ 1065^ 12597-'1 12606 s

9440s a
31 11760 *, - - - 20802- 23580- - - - - - - - - 32SS()Ji -
E AOAC

11080^ 2720 s 13720 s 3480 s 24505 16420S. 11300^

12630s'-! 3280 s 11700s- 3080s 2180s 15870- 1205Or • " • ^

1285s'-1- 11830- 14100-

34- 350 3500 2200 2800 400 800 350 900 250 700 350 700 1800 350

39 9708 9287 s
7146 2719 12675 3074 - 1337 14950 101 42 66 130
9067 6253^ 117* ND ND 7 12529 107
2429 12065 3127 1139 14227 102 61 53 144 146- ND 21 12137 107
Ji- -12"
1. 60, No. 6,1977

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 h h 14900h . 3700h 35300 <300 <300 <S00 <300 i3£00 <300 «
41 oUU
1145Og 10950S-4 1470O5'-1- 3520 s 197006 _65£'-£ -35£ -105-'•= -8Osfl270Og -lOO^f §
s s 6 s s
s 8
10500- 1125d£'J- 157OO£'4- 356QS 19200 -60 "- -.50 s -180 £ > - a -35 540 '* • ^^ -160 - >
10150- 79OO£'l 9700 s '- 1 - 12000s ^

10960 20 340 1370 243 158 238 148 3 63 170 160

12380 0 280 1700 238 163 213 108 10 53 550 138

" All data are on an "as received" sample basis. Designations of data, by way of (example) are as follows: reported by collaborator (1000); calculated by Associate
Referees (1000°); reported by collaborator and considered for statistical analysis (1000); calculated by Associate Referees and considered for statistical analysis (1000s);
reported by collaborator and used for calculations (1000);calculated by Associate Referees and used for calculations (1000f)- For reported data, calculations were checked
using the ng, ng/ml, solution volumes, and sample weights reported, to correct some obvious arithmetic errors, and data have been rounded-off to, at best, 1 ng/g; a
an extra digit was extracted by the Associate Referees in some cases of excessive rounding-off. The number of digits is unrelated to precision. Each result reflects 2
one determination unless otherwise indicated. Appropriate reagent blank corrections were not always made by collaborators. D
6
Shortages of some samples resulted in insufficient a mounts being sent to some laboratories for duplicate analyses requested. This laboratory ascertained by visual H
inspection and consultation with one of the Associate Referees that Samples 1, 8, and 9 were NBS Standard Reference Materials. These samples were available in 02
Laboratory 1 and were used to perform duplicates I2, 82, and 92.
" Not determined or not calculated.
d
Reported as not detectable. 3
" Results were calculated by the Associate Referees; see text.
1
All results from Laboratories 3, 34, and 49 were excluded from statistical calculations because most results from these laboratories appeared to have large system-
atic deviations; in addition, 5 results of Laboratory 3 are in the "less than . . ." form and unusable. Other "less than . . ." data from other laboratories were also
excluded. The 5 results from Collaborators 28, 29, and 41 for Sample 5B were excluded because of erroneous sample handling
" Results excluded from statistical analysis as outliers because the laboratory mean for the sample deviates excessively from the mean over all laboratories.
* Mean of 2 determinations.
1
The 42 ng/g reported appeared in error; recalculation gave 118 ng/g based on the 2.36 ng/ml reported.
' Results excluded from statistical analysis as outliers because within-digestion variance appeared too large relative to that for the other laboratories.
* Results were rounded-off to 2 figures by Collaborator 15 who reports "we have reported our values well beyond their significance." An additional figure was obtained
by the Associate Referee from the reported ng/ml; values have been rounded-off to 3 figures or 5 ng/g.
' Value of 6030 ng/g reported by Collaborator 15 seems in error due to erroneous value for volume used in calculation: 12070 ng/g was recalculated by Associate
Referee from the 1216 and 599 ng reported and 10 and 5 ml volumes, respectively, obtained from recorder charts.
™ Second digestion of Sample 4.
" Calculated by Associate Referee from ng/ml reported on recorder charts. Data obtained at both low and high scale expansion are listed although some data at low §
scale expansion are negative and inaccurate.
0
Calculated by Associate Referee from reported ng/ml and sample weights; differ slightly from reported values of 8800, 2700, 11100, and 14 ng/g, respectively.
p
Collaborator24 remarked that high reagent blanksand irreproducible calibration curves led to inconsistent and inaccurate results and suggested that these results
for arsenic not be considered.
" 5 ml of solution in ampoule D added instead of 5 ml of 6.25-fold diluted solution as instructed.
r
Average standard curves were used because a sufficient number of standards were not always included in each run.
8
Solution D was not added as instructed.
' Laboratory 39 knew identity of Sample 9 and used NBS Spinach for determination 92.
" 4 ml of solution in ampoule D added instead of diluted solution as instructed.

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 Table 3. Collaborative sample results (ng Se/g) for the hydride AAS determination of selenium in foods*
Sample/Solution

58 57b 4115 3894 1868 2700 ND^ -£- U98 107 1000 1034*" 8 292 105 684 1946 125
73 2i 4195 3595 2067 2839 ND 1398 102 1150 1064^ 25 263 95 764 1936 117
-245

3- <50O - 2000 - 990 1100

4 3470 3620 1700 2530 1320 li 938 890 J>4 194 125 563 1650 195
J64(lj)2 3510 893 54 T68 565 1650

las
3530 1590 2470 55J 1120 131 890 125 139

5 ND 187*1 1696^ S00-? ND e ND ND 20\

-11s 48s 135-i 195- -12O|>£
-T55 IP 1755--8 'iMeS.t Jss 171- -330^'^ _4755.£ -Iio5.£ '•US'* -llO^'* 3?
s!> 310 - 5160 - 1820 3170 270 1070 915 60 5J) 525 2020 245

10 22 - 37U 3800 1900 2777 0 1335 147 962 68 265 40 687 1965 170
4021 3710 1920 2777 "1460 127 935 180 440 57_ 725 1900 80

11 190 - 4176 - 1540 2946^ M 1149 202 973 829 223| 379i 38 737 973 128
278 3597 1413 12541 1532^ 202 544 725 344* II7J- 54 658 1139

12 86 3548 832 2443 1159 180 491 132 80 703 590 121
- - 21
178 3760 687 2502 1149 193 539 173 123 672 720 166
32
13 ND 3200 2650 1800 2400 120 950 45 200 100 1600 - =-1
40 2900 TWO" 2400 110 7B" "67~5 1500
-3s
I -35 §
jp.
675 2100 175 3!
14 125 3600 3650 1950 2750 50 1500 200 1000 25 230 100
150 - 3600 3450 1850 2700 0 1270 150 900 10 140 125 575 2150 225
2800 1300 775 tr1
2350 1200 850

15* ND soi 39M& 203 Oil 2950- iooh 796!! ieit£ 3

373O| 30055 16255 n-h •JI
-375Ij 1365^ 405'i 10235 5955 -1085'i 175 s '-'- 6835 U2<fis -63s
3325^ 3945- 205Q5 28755 17755 1685.1 97OE 38S-S 555.1 778$ 1940
t -155.1
-i 2475^ I
3658(42)

16 - 4300^ - 800* ND 3600i ssoi ND 900± 1700^ 200^

51205 690" 1815| -3705-1 158Q5 5035 9481 3555 225| 1445 s 183|
55s 42805 10055 2530- -205.1 "1570s 8485 25|5 850s 1455 s 1755
65Q5

247^ 357-} 3400 370O* 300 2000 94 124 158 0 97 167 - 594 500 119
307i 2871 75BB 3400l 355 2200 79 179 154 87 87 257 559 300 119
400i 30(4 M 0 0 4800l 400 2300 200J 84 150 200 7 27 300 650 500 104
3001 30(H 150 2100 3 154 150 150 117 57 300 "550 $2-
42 °?J
1070^- 540 - - 290*s1 2200^ 131-
22895 9025 11745 05 10785 1405 5455 255 605 705 285 1236f
B4&5 8705 12145 05 U085 5605 30 s 605 55E 3155

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 3730S- 21302 2400R 12S0± 12S± lllSfi-s
635 40385 37585 2289 s 24535 195 1345 s 120 s 1178 30* 202^ 774~ 2065^ 97^
75S 4135 s 3625 s 1971 s 23085 3T-5 11055 T265 10435 3l« 1827i
38285 21745 -75 1215 I? 2062- 99 s
-3
2130 - 2030 2240 <100 1900 150 445 250 70 720 1680 140
250 1930 1970 2430 130 1260 TfO 620 <S0 75 665 1950 95
1055 2105 405 in
2305 -705 T5
ND 6100^ 7340^ 4539^- J849^ 164(& 24& 200h 270^-
- t
-125 49655.1 29505.1 4045 1950s 2080^'^ 1325^*1 287^'^ 2160s* fsS-4 H
725054 28055.4 53805.1 2375 JTJF 21605-4 200O5-1 2695-K •5625.1 28054 150054 3O7O S ' £ 365s*
5845 s ' 1 r?
25355.4 w
4580^ - SiflS eotfi 0s a
s s
58105 900 s 36005 0 990 •OS
5060= ?105 3350 s 4305 7755 54§5 S
4610 s "736s
a
2600 2920 880 2200 570 320 ND 640 1040 ND
2480 2920 760 420 320 520 , 92
1240(62)fl 27/5?/U22)A'
m °
1000(6 2)~'B 950(122)i.

160 25 25 1280 25

39 126 118 3872 3667 1894 2067 ND 186 142 1055 1030 116 83 686 2077 152 §
148 121 3583 3821 2023 2488 56 1447 162 T255 1136 IS 754 2003 107
3
41 4400^ - 29<Wg UOCfz JSSOf 1200% 300\ 700^ 10&
655 455 38505 208 OS 2730- 150- User 495 s ' 1 1570- 1380- lO^ 300 s 75^ 620 s 212Of 115 s
955 905 44705 21505 29005 T405 J46QS 590£'£ 14455 10305 1605.1 |55 735- 1360- T355

48 78 - 3425 - 1910 2605 27901 1445 2931 235 90 613 2190 148
a
35 4045 2045 2670 26951 1385 1181 145 103 605 1935 173 o
S!
»-* See Table 2.
' All results from Laboratories 3 and 34 were excluded from statistical calculation as most results from these laboratories appeared to have large systematic devia-
tions; in addition, 8 results of Laboratory 3 are in the "less than . . ." form and unusable. Additional data reported on duplicate digests 42, 62, and 122 were not con- 8
sidered.
™ Second or third digestions as indicated in parentheses.
" The 1240 ng/g reported by Collaborator 11 appeared in error; 1532 ng/g was recalculated from the 15.32 ng/ml reported.
' Calculated from ng/ml reported on recorder charts. Data obtained at both lowand high scale expansion are listed although some data at low scale expansion are
expected to be inaccurate.
» Some data as reported (4255 ng/g for 2i and 501 ng/g for 12) differed substantially from the check calculations recorded here.

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1424
excluded because most results from these labora-
tories appeared to have large systematic devia-
tions; in addition, 5 results for arsenic and 8
results for selenium reported by Collaborator 3
were in the unusable "less than . . ." form. All
arsenic results from Collaborator 49 were ex-
cluded on the basis of large systematic error. A
number of other results were also excluded be-
cause within-digestion variance appeared too
large relative to that for the other laboratories,
or the laboratory mean for the sample appeared
to deviate excessively from the mean over all
laboratories. (A subsequent paper in this series
(4) will give a more detailed discussion.)
The remaining sample data were subjected to
analysis of variance to arrive at estimates of
the between-laboratory standard deviation, sL,
between-digestion standard deviation, sA, and
within-digestion (between-determination) stand-
ard deviation, sD. Estimates of the precision
standard deviation, sR, and the overall or repro-
ducibility standard deviation, sx, were obtained
from the relations sR2 = sA2 + sD2, and Sx2 =
SL2 + SA2 + SD2- Standard errors, sx, of the mean
were computed from sx2 = [sL2 + sA2/nA + s D 2 /
wAnD]/nL, where nL is the number of labora-
tories contributing to the mean, nA is the num-
ber of independent analyses carried out on the
sample by each laboratory, and %> is the num-
ber of determinations on the digest. In this
study, the values nA = 1 or 2, and wD = 2 were
taken to represent the data fairly; the fact that
nA and %> were not consistent for all labora-
tories would have only a small effect on the
estimation of sx2 and was ignored. Statistical
results are summarized in Tables 4-7.
Tables 4 and 5 give mean concentrations of
arsenic and selenium found in the 13 samples,
together with the corresponding standard devia-
tions. Analyte concentrations used in most cal-
culations were not corrected for moisture con-
tent, the only exception being the data used for
accuracy comparisons in Tables 8-11. The inter-
pretation of each of the 5 standard deviations
shown is as follows: sL represents systematic
error among laboratories, sA reflects errors aris-
ing from the sample digestion procedure, sD is
the precision of the determination step, sR is the
precision of analysis expected in any one labor-
This report of the Associate Referees was presented at the
90th Annual Meeting of the AOAC, Oct. 18-21, 1976, at
Washington, DC.
JOURNAL OF THE AOAC (Vol. 60, N o . 6, 1977)
atory, and sx is the precision expected when anal-
yses are performed by different laboratories. For
the 6 samples analyzed in duplicate, it was pos-
sible to arrive at estimates of all 5 standard
deviations. Analytical data for the 7 samples
analyzed once by each collaborator furnished
only 2 of the standard deviations, sD and sx, and
the sum, V * L 2 + SA2- In the 12 cases (6 for
each element) for which sh, sA, and sD are avail-
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able, usually sL > sD > sA. This observation
points to some imprecision in the determination
step, most likely arising from the hydride gen-
eration-transport phases rather than the actual
atomic absorption measurement. The between-
digestion variance sA2 was the lowest of all 3
variances, indicating that the procedure of sample
digestion contributes little variation to the final
result. In fact, sA2 was negative at times (and
sA imaginary), suggesting that its real value was
a very small positive number. The largest stand-
ard deviation, sL shows the existence of labora-
tory-dependent systematic error in the method.
Coefficients of variation (CV) listed in Tables
6 and 7 also reflect the performance of the
tested method. The CV values based on sx are
fairly large for both elements and all samples,
indicating a wide and unacceptable range of sys-
tematic errors in the method. Coefficients based
on sR are smaller in comparison, but still seem
too high, even for samples containing /xg/g
amounts of analyte.
We attempted to establish the accuracy3 of
the method by comparing the analytical data
with levels for the samples obtained by inde-
pendent analyses using methods other than hy-
dride generation AAS. Tables 8 and 9 detail
results from reference analyses of the collabora-
tive samples, together with other data available
from the literature and other sources. A total of
6 different analytical methods (4 for arsenic and
3 for selenium), instrumental and destructive
neutron activation, isotope dilution spark source
mass spectrometry, fluorometry, light absorption
photometry, and graphite furnace AAS, were
used to measure concentrations of arsenic and
selenium. The lack of biological standard refer-
ence materials (only 2 NBS SRMs, Orchard
Leaves and Bovine Liver, are at present certi-
fied for selenium, and only 2, Orchard Leaves
3
Accuracy in this work is taken to mean the extent of
agreement of the method mean over all laboratories with a
reference value.
IHNAT & MILLER: HYDRIDE AAS METHOD FOR AS AND SE IN FOOD 1425

Table 4. Statistical evaluation of collaborative results for arsenic (ng/g)

Between-lab. Between-dig. Within-dig. Precision Reprod.

Sample Av. concn s t d dev ., s t d dev., std dev., s t d dev., s t d dev.,
(code No.) found" SL SA SD SH Sx

Skim milk powder (11) 0.12 (14) 24.9* 10.4 27.0

IAEA flour (10) 31.2 (14) 49.7h 32.0 59.1
Flour (12) 31.4 (15) 60.2* 26.6 b
65.8
NBS Bovine Liver (8) 43.3 (17) 25.9 22.8 17.6 27.0 37.4
Kale (7/14) 93.1 (17) 26.2 23.4 11.4 26.0 37.0
NBS Spinach (9) 114.0 (16) 45.5 c 23.1 20.3 49.8
239.3b _»

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Apple (5A) 917.9 (10) 58.9 246.5
Apple (5B) 1449 (3) 680.3b 147.5 _J> 696.1
Swordfish (4) 2377 (19) 1114.7s 230.1 -J> 1138.2
NBS Tuna (2) 3061 (19) 597.7 ' 397.8 337.6 686.4
NBS Orchard Leaves (1) 9265 (16) 184.7 569.0 600.0 826.9 847.3
IAEA oyster (3/13) 11447 (18) 1298.1 c 992.8 912.3 1586.6
Flounder (6) 13149 (15) 4265.2* 736.2 _& 4328.2

" Unconnected for moisture c o n t e n t ; n u m b e r of laboratories contributing t o t h e average is indicated in paren-

theses.
b
SL, SAand sa cannot be calculated; value between t h e s t a n d SA columns is V S L 2 + SA 2 .
c
Variance is negative; standard deviation is imaginary.

Table 5. Statistical evaluation of collaborative results for selenium (ng/g)

Between-lab. Between-dig. Within-dig. Precision Reprod.

Sample Av. concn std dev., std dev., std dev., std dev., std dev.,
(code No.) found" SL SA SD SH Si
Apple (5A) 31.0(10) 79.5b 30.9 —* 85.3
NBS Spinach (9) 42.9(12) 26.4 —" 34.4 29.4 39.5
NBS Orchard Leaves (1) 78.0(19) 53.3 40.3 35.7 53.8 75.7
Skim milk powder (11) 82.9(14) 23.2b 12.0 —h 26.1
Kale (7/14) 131.5(16) 40.9 13.1 23.5 26.9 49.0
Apple (5B) 160.3(4) 104.5b 77.9 —* 130.4
IAEA flour (10) 213.1(18) 83.0* 64.3 —b 104.9
Flour (12) 686.9(18) 68.5b 71.5 —* 99.0
NBS Bovine Liver (8) 774.5(19) 348.5 107.3 125.8 165.4 385.7
Flounder (6) 1319 (18) 232.46 156.0 —b 279.9
IAEA oyster (3/13) 1447 (19) 596.5 179.6 175.2 250.9 647.1
Swordfish (4) 2538 (17) 325.7* 189.4 —b 376.8
NBS Tuna (2) 3625 (19) 689.9 —c 293.3 276.9 743.4
b c
"• - See Table 4.

Table 6. Coefficients of variation for arsenic Table 7. Coefficients of variation for selenium

Av. concn. C o e f f . o f v a r ._
Av. concn Coeff. of var.,
Sample found, Sample found,
(code No.) ng/g" (code No.) ng/g" SR sx
Skim milk powder Apple (5A) 31.0 275.2
(11) 0.12 22500 NBS Spinach (9) 92.1
42.9 68.5
IAEA flour (10) 31.2 189.4 NBS Orchard
Flour (12) 31.4 209.6 Leaves (1) 69.0 97.1
78.0
NBS Bovine Liver Skim milk powder
(8) 43.3 62.4 86.4
(11) 82.9 31.5
Kale (7/14) 93.1 27.9 39.7
Kale (7/14) 131.5 20.5 37.3
NBS Spinach (9) 114.0 17.8 43.7
81.3
917.9 26.9
Apple (5B) 160.3
Apple (5A) 49.2
1449 IAEA flour (10) 213.
Apple (5B) 48.0 14.4
2377 Flour (12) 686.
Swordfish (4) 47.9
3061 NBS Bovine Liver
NBS Tuna (2) 11.0 22.4
(8) 774. 21.4 49.8
NBS Orchard
Flounder (6) 1319 21.2
Leaves (1) 9265 8.9 9.1 IAEA oyster (3/13) 1447 17.3 44.7
IAEA oyster (3/13) 11447 8.0 13.9 Swordfish (4) 2538 14.8
Flounder (6) 13149 32.9 NBS Tuna (2) 3625 7.6 20.5
" Uncorrected for moisture content.
b
SH cannot be calculated for these samples. «•' See Table 6.
 Table 8. Reference analyses (ng/g) of collaborative samples for arsenic9
Method and laboratory

NAA with cheim. Light absorption Light absorption Light absorption Graphite
b Other values
sepn— IDSSMS^- photometry with photometry with phototnetrv with furnace AAS&
(n) (n) ^ (n) AgDDC^ AgDDC^ (n)
Sample
code

10900 (1) 10000+2000 1 : 1060O+800(ll)i; 11000+1000(12)-;

9000(13)i; 13000(13)E

2 1100+100 (2) 1500+300 (S) 4200+500 (S) 1200 3200 (1) 1740 4600+300(11)-; 3600+200(12)°; 3300+200(12)-
4600+300(1 §
3)
3/13 6200+300 (2) 17000+900 (3) 8700 1290±200 (2) 10700 >
4 700+150 (2) 1900+300 (i) 7600 2700+100 (2) 1640 3360+4O3- o
6 5900+300 (2) 6500+1000 (2) 19400+1000^ (6) 7700 11800+200 (2) 9370 157OO+25O3-

>
a
7/14 100+20 (2) <50 260 127+29(16)-; 141 (range 110-220) (17,18)-

5^-; 49+6(12)- o
9 90+20 (2) <iooo 150+50^; 198+10^

10 10+5 (2) <iooo 50 24+8(19)-

p
11 <5 (2) <50

12 <3 (2) <50

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» «•-»
 ° All concentrations, initially reported on an as-received basis, have been corrected to a dry sample basis; all "other values" are also presumably on a dry basis.
Mean concentrations ± standard error of n analyses/determinations are listed. Values chosen for the calculation of selected reference levels are underscored.
6
P. Schramel; neutron activation analysis with radiochemical separation after irradiation (6).
" J. A. Carter and D. L. Donohue; low temperature ashing, isotope dilution spark source mass spectrometry (7), using 11000 ng As/g of NBS Orchard Leaves SRM 1571 8-
as reference value.
d
M. Stoeppler and W. Matthes; digestion with HNO3-HCIO4-H2SO4 as described for this collaborative study, and photometry using AgDDC (8, 9).
' E. Sandi and M. T. Lo; ashing at 480°C with Mg(NO3>2, and photometry using AgDDC according to modified AOAC method (10).
1
K. H. Tarn; ashing at 500°C with Mg(NO 3 ) 2 and MgO, and photometry using AgDDC.
' R. B. Flemming and H. C. Freeman; digestion with HNO3, determination using Perkin-Elmer HGA-2000 graphite furnace.
* Concentration (ng/g) ± uncertainty as defined in footnotes; numbers in parentheses refer to references at end of paper.
' NBS certified value ± twice standard deviation or entire range of observed results. o
' Mean ± standard deviation of >4 determinations using neutron activation analysis with chemical separation after irradiation. 3
* Mean ± standard deviation based on analysis of 3 replicate 1 g samples using hydride AAS method. a
1 a
Dry ashing/hydride AAS method following procedures of Dalton and Malanoski (14, 15); no uncertainty stated.
m
Value obtained by another laboratory using an unspecified AAS method.
" N B S informational value, mean ± standard deviation of analyses of 8 samples, using neutron activation analysis with chemical separation.
° FDA instrumental neutron activation value; uncertainty includes counting statistics.
"Sample still under analysis in IAEA intercomparison program; IAEA value is not yet available. n
* H. J. Miller; hydride AAS; mean ± standard error. a
' Sample 6 was sent to this laboratory under 2 different codes. o
* Best mean ± standard deviation; 16 activation analyses by 3 different laboratories.
o
1
Best value and range of laboratory means; 27 activation analyses by 7 laboratories.
" Uncertified NBS value.
" IAEA value ± t X standard error for 95% confidence limits based on 10 analyses by 3 laboratories using neutron activation analysis and light absorption photo-
metry. Value reported by IAEA (19) corrected to dry basis.
O
00
H

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 to
00

Table 9. Reference analyses <ng/g) of collaborative samples for selenium"

Method and laboratory

NAA with chem. INAA~ (n) INAA- (n) t u v w

sepnS- (n) Fluorometry— Fluorometry Fluorometry" Fluorometry— Other values
Sample (n) (n) (n) (n)
code

95+3 (3) 79+2 (4) 80+10^ 80+10(ll)J-; 55+9(12)-; 118+17-;

Tr<5)^ ~
2 23900+2100 (2) 3850+120 (2) 4430+210 (3) 4180+50 (S) 3710+60 (4) 3710+130 (2) 3300+300(11)-; 3600+100(12)-; 3720+1705-

3/13 1200+210 (2) 1830+150 (2) 2140+210 (3) 2230+40 (5) 2010+50 (6) 1880+50 (2) 2220+30 (6)

4 1100+200 (2) 2900+60 (2) 3270+200 (3) 2950+40 (S) 2830+70 (6) 2360+170 (4) 2900+50 (2) 2910+45(5)i

5A 5+1 (4) 66+16^; 23+2^

5B 6+3 (4) 1+1 (2) 203+93.

6 660+150 (2) 1430+70 (2) 1630+200 (3) H40+40 (5) 1400+30 tb) 1420+180 (4) 1470+15 (4) 1450+50^; 1580+203- W
7/14 140+15 12) <2OO+2OO (2) <200 (3) 125+4 (S) 135+2 (6) 122+10 (3) 126+2 (6) 200+52-; 148+14(16)—; 15O?(17)—;
121?140 (range 20-150) (18)-££- I
1260+40 (S) 1220+5 (4) UOQ+100^-; 1090+50(11)^-; 980+10(12)-;
+^0^; 1210+30(5)2.

9 <100 (2) <30O+300 (2) <200 (3) 45+10 (4) 38+1 (6)

10 370+60 (2) 260+20 (2) 270+40 (3) 295+10 (S) 280+4 (6) 235+20 (4) 286+5 (4) 187+34-
p
11 <30 (2) <200+200 (2) <100 (3) 97+2 IS) 100+2 IB) 85+3 (2) 99+1 f4J 112+6(5)^

12 630+110 (2) 750+30 (2) 790+40 (3) 860+10 (4) 805+15 (6) 830+10 (2) 850+15 14) 804+12(5)^

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^ i
 « .r

°-« See Table 8.

r
J. F. Emery; instrumental neutron activation analysis. a
* J. T. Tanner and M. H. Friedman; instrumental neutron activation analysis (20, 21).
' R. A. Cloutier; fluorometry using 2.3-diaminonaphthalene and AOAC method 25.117-25.120. The same reagent was used by all other analysts using fluorometry.
* J. H. Watkinson and M.T. Wingerden; digestion with HNO3-HCIO4, determination using semiautomated adaptation of fluorometric method of Watkinson (22).
* A. Sosner and R. Cohen; fluorometry, AOAC method 25.117-25.120 with omission of H2O2.
" J. Jago; digestion with HNO3-HCIO4, fluorometry according to method of N. L. Wilson (23), essentially the method of Watkinson (22). f
* M. Ihnat; hydride AAS; mean ± standard error.
'Mean ± standard error; based on 14 (Sam pie 1), 16 (Samples 4, 8, and 12), and 18 (Sample 11) analyses by as many laboratories during collaborative study of f luoro- r°
metric method, 25.117-25.120 (5). a
' Sample still under analysis in IAEA intercomparison program; IAEA value is not yet available; the listed value was determined by M. Ihnat (seex). o
"" Mean ± standard deviation based on 20 analyses by 4 laboratories using fluorometry and activation analysis. S
66
Best value. Agreement between results from activation analysis (ranges of means from 4 laboratories, 17-620 ng/g) and fluorometry (individual means from 2 labo- 3
ratories, 140 and 140 ng/g) is poor. H
" Best mean, most probable value, and range, respectively. Bowen writes " . . . for kale the analyses show a bimodal distribution with two 'most probable' values
around 40 and 140 ng/g. Further work is needed to establish which of these is correct but it is suspected that the volatility of the element may have given rise to the CO

lower values."
dd
NBS certified value with 95% confidence limits estimate of uncertainty.
O
d

GO

Z
u

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1430 JOURNAL OF THE AOAC (Vol. 60, N o . 6, 1977)

Table 10. Accuracy of the hydride AAS method 11 as well as those in Tables 8 and 9 have been
for arsenic" corrected for sample moisture contents. Com-
Selected ref., Av. found, parisons for arsenic (Table 10) could only be
Sample ng/g 6 ng/g" made for 6 of the 13 samples for the reason
IAEA flour 24 ±8 (3)"1 35 ±17 (14)
given above. Agreement was excellent between
NBS Bovine Liver 55 (NBS) 6
46 ±8 (17) the hydride and NBS values for Orchard Leaves.
Bowen's kale 141 ±15 (7) 101 ±9 (17) Remarkedly good agreement was obtained for
NBS Spinach 150±5O(NBSy 120 ±13 (16)
NBS Tuna 3900 ±300 (4) 3160 ±145 (19)
IAEA flour, NBS Bovine Liver, kale, and NBS
NBS Orchard Leaves 10000±2000 (NBS)/ 9940 ±145 (16) Spinach in spite of the low arsenic levels present

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in these examples; agreement was poorer for
" All results are on a dry sample basis and are re- NBS Tuna. For 5 samples (except IAEA flour),
ported as mean ± standard error (number of labora-
tories), unless otherwise noted. however, hydride AAS values were consistently
b
Reference levels were calculated by averaging the lower.
selected data of Table 8 giving identical weights to the
mean results from each laboratory. Several samples For selenium (Table 11), some sort of refer-
are not listed because agreement among reference ence numbers were available for all 13 samples.
laboratories was poor, making it impossible to select
data. For 8 samples, NBS Spinach, NBS Orchard
c
Standard error, sx is defined in text. Leaves, skim milk powder, kale, IAEA flour,
d
Uncertainty reported by IAEA is ± t X standard flour, flounder, and NBS Tuna, the accuracy of
error for 95% confidence level.
* Tentative NBS level. the hydride AAS method was very good. Hy-
1
NBS certified concentration including twice the dride AAS results for NBS Bovine Liver, IAEA
standard deviation or the entire range of observed oyster, and swordfish were, respectively, 25, 24,
results.
and 11% lower than selected reference levels.
Firm comparisons between data were not made
and Spinach, are certified for arsenic) made for the 2 apple samples because only limited
these analyses, together with other reported val- data were available from reference laboratories,
ues, invaluable (at least for selenium) in arriv- and collaborators analyzed spiked samples,
ing at selected reference estimates of levels in a whereas reference laboratories analyzed unspiked
number of samples. samples. The 3 samples exhibiting low recoveries
Only levels generated by methods other than are either meat or fish, suggesting a systematic
hydride AAS formed the basis for selection; sample matrix effect either in hydride evolution
non-collaborative results obtained by hydride or sample treatment. Curiously, tuna, a sample
generation AAS are included in Tables 8 and 9 also of marine origin as oyster and swordfish,
only for information. Seemingly acceptable val- showed good agreement of determined selenium
ues were chosen by inspection and pooled to levels. Average selenium levels found in the
give mean selected levels shown in Tables 10 and 1973 collaborative study of a fluorometric method
11. For NBS Orchard Leaves, Bovine Liver, and for selenium (5) are also included in Table 11
Spinach, the NBS certified levels were used for information. Comparison of these data with
where available. Large disagreement among ref- either NBS values or other appropriately selected
erence results for arsenic made selection difficult reference levels (excluding the same fluorometric
or impossible with the consequence that very data) demonstrates the good accuracy of fluoro-
few reference results are available for this ele- metric analysis. These tests for accuracy indi-
ment ; its determination seems difficult. For sele- cate that for both elements, accuracy of the
nium, on the other hand, between-laboratory hydride AAS method is quite acceptable for a
and between-method agreement is excellent in number of different (but not all) sample types,
comparison, and many data were available for in spite of the presence of a large variance asso-
estimating selenium levels. ciated with systematic error.
Comparisons of arsenic and selenium concen- In arriving at the foregoing observations of
trations obtained with the hydride AAS method, method performance, it must be kept in mind
with selected reference levels, are presented in that the data were screened and a number of
Tables 10 and 11. Concentrations listed here do outlying observations have been rejected prior
not agree with the corresponding ones in Tables to final calculations. Whereas the rejection of
4-7 because the concentrations in Tables 10 and some results seemed obvious, the perpetual ques-
IHNAT & MILLER: HYDRIDE AAS METHOD FOR AS AND SE IN FOOD 1431

Table 11. Accuracy of the hydride AAS method for selenium0

Fluorometric
Selected ref., Av. found, method (5),
Sample ng/g 6 ng/g c ng/g

Mclntosh apple 5 ± 1 ay 32 ±27 doy

Delicious apple 4 ±4 (2)" 170 ±63
NBS Spinach 42 ±4 (2) 45 ±9 (12)
w
NBS Orchard Leaves 80 ±10 (NBS)/ 84±15 (19) 92 ±5 (14)
Skim milk powder 109 ±5 (22) 87 ±7 (14) 112 ±6 (18)
[95±3 (4)]»

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Bowen's kale 135 ±3 (8) 143 ±12 (16)
IAEA flour 271 ±9 (6) 239 ±25 (18)
Flour 807 ±11 (22) 760 ±22 (18) 804±12 (16)
[814 ±17 (6)]'
NBS Bovine Liver 1100 ±100 (NBSy 826 ±89 (19) 1200 ±30 (16)
Flounder 1470 ±34 (6) 1350 ±62 (18)
IAEA oyster 2050 ±160 (6) 1550 ±150 (19)
Swordfish 2900 ±46 (22) 2590 ±87 (17) 2910 ±45 (16)
[2870 ±120 (6)]»
NBS Tuna 3860 ±160 (6) 3740 ±170 (19)

• All results are on a dry sample basis and are reported as mean ± standard error (number of laboratories),
unless otherwise noted.
b
Reference levels were calculated by averaging the selected data of Table 9 giving identical weights to the mean
(or only) results from each laboratory. Two low values of 17and 46 ng/g and one high value of 620 ng/g determined
by activation analysis for kale (17) were rejected.
e
Standard error s i is defined in text.
d
Insufficient data.
' These samples were fortified by collaborators using solution from ampoule D whereas reference analyses were
done on non-fortified samples.
1
NBS certified concentration including 95% confidence limits or entire range of observed results.
' Reference levels calculated by omitting data from collaborative study of fluorometric method (5).

tion arises as to whether rejected values were
bona fide members of the data population or
aberrant data. Some results such as the 0 ng
As/g reported by Collaborator 49 for several
samples are likely to be aberrant values but
could conceivably arise from, and flag a weak
point in the method; the assessment of method
performance characteristics should be tempered
by this behavior and an attempt should be made
to locate the cause of the problem. The method
cannot be recommended if some explanation
cannot be found for the large numbers of out-
lying observations. Whether in fact the method
or laboratory-related techniques are responsible
for the observed performance is a question to be
tackled in the detailed treatment of data pre-
sented in the following reports (3, 4).
In summary, this study has shown that under
conditions of a specified sample treatment pro-
cedure and essentially free choice of hydride
generation equipment and technique, the per-
formance of an acid-digestion NaBH4-facilitated
hydride generation AAS method for estimating
arsenic and selenium in foods is fairly good with
respect to accuracy for many, but not all, food
samples tested and unacceptable with respect to
precision. Recommendations are contained in the
following report.
Acknowledgments
The Associate Referees are deeply grateful to
the following for their efforts in contributing to
the study as collaborators and reference labo-
ratories :
R. C. Abel, Food and Drug Administration,
Baltimore, MD
F. A. J. Armstrong and A. Lutz, Environment
Canada, Freshwater Institute, Winnipeg, Mani-
toba, Canada
R. Arreola, R. R. Cortez, D. F. Fischer, and
W. R. Holtz, Joint Water Pollution Control
Plant, Water Quality Laboratory, Carson, CA
E. E. Bailey, and J. P. Minyard, Jr, Missis-
sippi State Chemical Laboratory, Mississippi
State, MS
Z. Baraniak, B. J. Houlahan, and D. Kova-
chic, Agriculture Canada, Plant Products Divi-
sion, Ottawa, Ontario, Canada
R. A. Buckley and G. B. Jones, CSIRO, Divi-
sion of Human Nutrition, Adelaide, Australia
S. G. Capar and J. W. Jones, Food and Drug
Administration, Washington, DC
1432
J. A. Carter, D. L. Donohue, and J. F. Emery,
Oak Ridge National Laboratory, Analytical
Chemistry Division, Oak Ridge, TN
C. Y. Chan and P. N. Vijan, Ontario Min-
istry of the Environment, Air Quality Labora-
tory, Toronto, Ontario, Canada
G. D. Christian and E. J. Knudson, Depart-
ment of Chemistry, S. Olsen and W. R. Schell,
Laboratory of Radiation Ecology, University of
Washington, Seattle, WA
0. E. Clinton, J. H. Watkinson, and M. T.
Wingerden, New Zealand Ministry of Agricul-
ture and Fisheries, Ruakura Agricultural Re-
search Centre, Hamilton, New Zealand
R. A. Cloutier, Agriculture Canada, Chemistry
and Biology Research Institute, Ottawa, On-
tario, Canada
R. Cohen and A. Sosner, Folkman and Dr.
Koffler Ltd., Tel Aviv, Israel
T. T. Collins, M. A. Nevelle, T. Surtes, and
J. R. Tuschall, Jr, CDM/Limnetics, Milwaukee,
WI
E. J. Dixon, Laboratory of the Government
Chemist, Department of Industry, London, UK
R. J. Emerick and 0. E. Olson, South Dakota
State University, Department of Chemistry,
Brookings, SD
F. E. Estiandan, M. Shimabukuro, and N.
Uichanco, Carnation Research Laboratories, Van
Nuys, CA
F. J. Fernandez, The Perkin-Elmer Corp.,
Norwalk, CT
R. B. Flemming and H. C. Freeman, Environ-
ment Canada, Fisheries and Marine Service,
Halifax, Nova Scotia, Canada-
M. H. Friedman and J. T. Tanner, Food and
Drug Administration, Washington, DC
L. R. Hageman, State of Montana Depart-
ment of Agriculture, Montana State University,
Bozeman, MT
M. B. Hocking and M. Ko, University of Vic-
toria, Department of Chemistry, Victoria, Brit-
ish Columbia, Canada
Y. Inoue, M. Munemori, and T. Nakahara,
University of Osaka Prefecture, College of Engi-
neering, Mozu-umemachi, Saki 591, Japan
R. A. Isaac and W. C. Johnson, University of
Georgia, Soil Testing and Plant Analysis Labo-
ratory, Athens, GA
J. Jago, Government Chemical Laboratories,
Perth, Australia
M. T. Lo, E. Sandi, and K. H. Tam, Health
JOURNAL or THE AOAC (Vol. 60, No. 6,1977)
and Welfare Canada, Food Directorate, Ottawa,
Ontario, Canada
B. Loescher, Ontario Ministry of the Environ-
ment, Laboratory Services Branch, Rexdale, On-
tario, Canada
W. Matthes and M. Stoeppler, Nuclear Re-
search Centre, Institute of Chemistry, 4 Jue-
lich, West Germany
R. W. Marts, Food and Drug Administration,
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Kansas City, MO
C. M. McGregor and J. Parker-Sutton, Min-
istry of Agriculture, Fisheries, and Food, Derby,
UK
J. C. M. Mol-Krijnen, B. E. Murphy, and
S. R. B. Solly, New Zealand Ministry of Agri-
culture and Fisheries, Wallaceville Animal Re-
search Centre, Upper Hutt, New Zealand
S. M. Muir and F. J. Schmidt, Illinois Envi-
ronmental Protection Agency, Champaign, IL
W. Pfannhauser and H. Woidich, Forschungs-
institut der Ernahrungswirtschaft, Vienna,
Austria
R. V. D. Robert, National Institute for Metal-
lurgy, Johannesburg, South Africa
P. Schramel, Gesellschaft fur Strahlen-und
Umweltforschung mbH., Neuherberg/Munchen,
West Germany
A. M. Sulek, National Canners Association,
Washington, DC
J. R. Williams, Food and Drug Administra-
tion, San Francisco, CA
R. J. Westerby, Agriculture Canada, Chem-
istry and Biology Research Institute, Ottawa,
Ontario, Canada
The authors are indebted to the following who
supplied on short notice most of the samples
used in the study: H. J. M. Bowen, Department
of Chemistry, The University, Reading, UK
(kale); International Atomic Energy Agency,
International Laboratory of Marine Radioac-
tivity, Monaco-ville, Monaco (oyster), Analyti-
cal Quality Control Services, Vienna, Austria
(flour); U.S. Food and Drug Administration,
Baltimore, MD (NBS Bovine Liver and Orchard
Leaves); J. T. Tanner, U.S. Food and Drug Ad-
ministration, Washington, DC (NBS Spinach
and Tuna); and J. Graham, Environment Can-
ada, Ottawa and K. Spencer, Environment Can-
ada, St. John's, Newfoundland (flounder). The
technical assistance of R. J. Westerby, Agricul-
ture Canada, Ottawa, R. C. Abel, U.S. Food and
Drug Administration, Baltimore, MD, and J.
IHNAT & MILLER: HYDRIDE AAS METHOD FOR AS AND SE IN FOOD
Jones, Agriculture Canada, Food Research Insti-
tute, Ottawa (for freeze-drying several fish sam-
ples) is appreciated. Moisture determinations
were performed by Analytical Chemistry Serv-
ices, Chemistry and Biology Research Institute,
Agriculture Canada. The assistance of B.
Thompson, Statistical Research Service, Agricul-
ture Canada, with the design of the study and
the statistical computations is gratefully ac-
knowledged.
REFERENCES
(1) Official Methods of Analysis (1975) 12th Ed.,
AOAC, Washington, DC
(2) Ihnat, M., & Miller, H. J. (1977) 60, 813-825
(3) Ihnat, M., & Thompson, B. (1977), in prepara-
tion
(4) Thompson, B., & Ihnat, M. (1977), in prepara-
tion
(5) Ihnat, M. (1974) JAOAC 57, 373-378
(6) Samsahl, K. (1967) Anal. Chem. 39, 1480-1483
(7) Carter, J. A., Donohue, D. L., Franklin, J. C ,
& Stelzner, R. W. (1975) 9th Annual Con-
ference on Trace Substances in Environ-
mental Health, University of Missouri, Co-
lumbia, MO, June 9-12, pp. 303-310
(8) Woidich, H., & Pfannhauser, W. (1975) Z.
Lebensm. Unters. Forsch. 159, 323-327
(9) Dahl, R. (1973) Int. Ber. der KFA Juelich
(unpublished).
(10) Official Methods of Analysis (1975) 12th Ed.,
AOAC, Washington, DC, sec. 25.012
(11) Orvini, E., Gillis, T. E., & LaFleur, P. D.
(1974) Anal. Chem. 46, 1294-1297
1433
(12) Fiorino, J. A., Jones, J. W., & Capar, S. G.
(1976) Anal. Chem. 48, 120-125
(13) Zook, E. G., Powell, J. J., Hackley, B. M.,
Emerson, J. A., Brooker, J. R., & Knobl,
G. M., Jr (1976) J. Agric. Food Chem. 24, 47-
53
(14) Dalton, E. F., & Malanoski, A. J. (1969)
JAOAC 52, 1035-1038
(15) Dalton, E. F., & Malanoski, A. J. (1971) At.
Absorp. Newsl. 10, 92-93
Downloaded from https://academic.oup.com/jaoac/article/60/6/1414/5710230 by guest on 04 November 2020
(16) Bowen, H. J. M. (1967) Analyst 92, 124-131
(17) Bowen, H. J. M. (1969) Adv. Act. Anal. 1,
101-113
(18) Bowen, H. J. M. (1974) J. Radioanal. Chem.
19, 215-226
(19) Gorski, L., Heinonen, J., & Suschny, O.
(1974) "Final Report on the Intercomparison
of Trace Multielement Analysis in Dried Ani-
mal Whole Blood (A-2), Calcinated Animal
Bone (A-3/1), Powdered Milk (AS), Wheat
Flour (V-2/1), and Dried Potatoes (V-4),"
International Atomic Energy Agency, Vienna,
Austria, No. IAEA/RL/25
(20) Friedman, M. H., & Tanner, J. T. (1975)
Trans. Am. Nucl. Soc. 21, 97-98
(21) Friedman, M. H., & Tanner, J. T. (1975) / .
Radioanal. Chem. 25, 269-273
(22) Watkinson, J. H. (1966) Anal. Chem. 38, 92-
97
(23) Wilson, N. L. (1969) "The Determination of
Selenium in Agricultural Materials," Report
of Investigations No. 6, Government Chemi-
cal Laboratories, Perth, Western Australia
Received December 15, 1976.